Our global objective is to develop a molecular understanding of epithelial stem cell lineages and morphogenesis in mammalian skin, and bring this research to a clinical setting. Our focus is on how skin stem ceils utilize cytoskeletal connections to intercellular junctions. Knowledge of the proteins involved in linking the actin filament (AF) cytoskeleton to adherens junctions (AJs), and elucidating how these connections are regulated are key to understanding how self-renewing skin epithelium maintains a surface barrier and how a stem cell can give rise to a hair follicle (HF). Elucidating how microtubules (MTs) link to AJs is important for understanding how the stratified epidermis maintains a single layer of dividing cells, and how spindle polarity changes when stem cells are activated to produce HFs. To examine cytoskeletal-junction dynamics during epidermal homeostasis and wound-healing, mice displaying fluorescently labeled AFs, MTs and AJs will be engineered, and used to derive three dimensional epidermal cultures. A molecular understanding of these dynamics will be obtained by focusing on two cytoskeletal-junction linking proteins that surfaced as being key from our prior studies: beta-catenin, which links AFs and AJs, and ACF7, which can bind MTs and link them to intercellular junctions. Through yeast two-hybrid screens, biochemical and molecular approaches, alpha-catenin and ACF7's interacting partners will be defined and characterized. Through mouse genetics, the functional significance of their varied associations will be ascertained. Finally, as the major players in these processes unfold, microarray analyses will be conducted, to define global changes in AJ-cytoskeletal gene expression that occur concomitantly as stem cells receive external cues to remodel their AJ-cytoskeletal connections and initiate HFs. Understanding how these cytoskeletal linkages are regulated in normal skin is a prerequisite to elucidating how defects in these processes lead to genetic disorders, including skin cancers. Past and present AR27883 research provides an excellent illustration of how molecular skin biology can help to generate new and improved tools for the diagnosis and treatment for human skin disease.